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Team:Tuebingen/Notebook/Protocols/control-restriction
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Latest revision as of 12:10, 4 October 2013
Control Restriction Digest
Reagents
| 1.5 µL | 10x NEBuffer 2 |
| 0.15 µL | 100x BSA |
| 3 µL | DNA from Mini-Prep (c > 50 ng/µL) |
| 0.15 µL | EcoRI or XbaI |
| 0.15 µL | PstI or SpeI |
| to 15 µL | Aqua dest. |
Procedure
- Around noon, mix all reagents in an 1.5 mL Eppendorf-tube and incubate at 37 °C until afternoon.
- Heat inactivate restriction enzymes at 80°C for 20 min.
- Add 1/10 volume of 10X Antarctic Phosphatase and mix. Incubate at 37 °C for 15 min for 5' extensions.
- Heat inactivate phosphatase at 70 °C for 5 min.
- Run control-gel in order to check a ligation / transformation.
